Thioredoxin reductase 1 (TxRd1), is an essential selenoprotein with defined roles in redox regulation of cell growth, death (apoptosis), transcription, DNA repair, and angiogenesis. TxRd1 is overexpressed in many human cancers and these elevated levels of TxRd1 have been associated with poor prognosis due to enhanced cellular resistance to chemo- and radiation therapy. Chemical inhibitors of TxRd have yielded promising results in preclinical and clinical trials as cancer therapeutics and radiation sensitizers. Curcumin is a naturally occurring plant polyphenolic antioxidant with potent chemopreventive and antintumorigenic properties in preclinical animal models and a demonstrated low toxicity profile in Phase I clinical trials Recent data demonstrate a covalent interaction between curcumin and the conserved c-terminal cys-x-sec-cys motif in TxRd1. resulting in inhibition ofthe catalytic interaction between TxRd and thioredoxin. The electrons donated by NADPH are instead diverted to O2, to form superoxide and ultimately hydrogen peroxide. In this proposal we will investigate the role of TxRd1 as a critical mediator of the antitumorigeniceffects of curcumin alone and in combination with ionizing radiation in oraopharyngeal cancer cell lines, in vitro and in vivo. . Our preliminary data demonstrated >50 fold higher level of TxRd1 in the human squamous carcinoma cell line FaDu compared to either an oral leukoplakia cell line MSK-Leuk1 or nontransformed keratinocytes. In specific aim 1, we plan to knockdown TxRd1 levels in FaDu cells using shRNA to the 3'UTR of the human TxRd1 gene cloned into the pSilencer expression vector. Conversely, TxRd1 will be overexpressed in MSK-leuk1 cells by transfection with the full length wild type TxRd1 gene, including the selenocysteine insertion element, from the 3'UTR. The stable clones selected above will be used to determine whether TxRd1 mediates the anti-proliferative and pro-apoptotic actions of curcumin, in vitro. In specific aim 2, we will use the stable clones selected above, to determine whether curcumin's efficacy as a radiation sensitizer also depends on inhibition of TxRd1. In specific aim 3, the TxRd1 knockdown and knock in cell lines, and the parental cell lines, will be grown as tumors in the cervical subcutaneous tissue of athymic nude mice. The efficacy of curcumin administered in the diet alone or in combination with ionizing radiation on orthotopic tumor growth and tumor cell death will be measured. Since curcumin is a natural bioactive food component with no overt systemic toxicities, promising results from these preclinical experiments can be readily translated into a chemoprevention schemes or combined chemo-radiation therapy protocol for treating head and neck cancers. PUBLIC HEALTH RELEVANCE: Head and neck cancers (HNC) are the sixth most common form of cancer worldwide, with over 500,000 new cases diagnosed annually. Although advances in surgery, and combined of radio and chemotherapies, have decreased morbidity, the survival rate at 5 years after initial prognosis remains at <50%. Moreover, since smoking and alcohol use are the two most prevalent risk factors associated with HNSCC, field cancerization leads to second primary cancers in 20% of HNC patients. Cucumin, a natural product derived from the root of the plant Curcuma longa, has shown promise as a cancer chemopreventive, chemotherapeutic and radiation sensitizer, in preclinical and phase 1 clinical trials. The primary drawback to the use of curcumin is limited bioavailability due to low concentrations of the drug reaching the circulation. However, curcumin has been effective in tumors, such as colon and HNC, where direct access to the drug is possible. The primary aim of this study is to develop curcumin as a chemotherapeutic and radiation sensitizer for treating HNC..